Method for treating hypercholesterolemia with polyallylamine polymers

ABSTRACT

A method for removing bile salts from a patient that includes administering to the patient a therapeutically effective amount of a non-absorbable amine polymers characterized by a repeat unit having the formula:  
                 
and salts thereof, where n is a positive integer and x is zero or an integer between 1 and about 4.

RELATED APPLICATIONS

This application is a Continuation of U.S. application Ser. No.10/158,207, filed May 29, 2002, which is a Continuation of U.S.application Ser. No. 08/979,096, filed Nov. 26, 1997, now U.S. Pat. No.6,423,754 which is a Continuation-in-Part of U.S. application Ser. No.:08/927,247, filed Sep. 11, 1997 now abandoned, which is a Continuationof U.S. application Ser. No.: 08/878,422, filed Jun. 18, 1997 nowabandoned. The entire teachings of the above applications areincorporated herein by reference.

BACKGROUND OF THE INVENTION

Reabsorption of bile acids from the intestine conserves lipoproteincholesterol in the bloodstream. Conversely, blood cholesterol levels canbe diminished by reducing reabsorption of bile acids.

One method of reducing the amount of bile acids that are reabsorbed and,thus, reducing serum cholesterol is the oral administration of compoundsthat sequester the bile acids and cannot themselves be absorbed. Thesequestered bile acids consequently are excreted.

Compounds which have been suggested for bile acid sequestration includevarious ion exchange polymers. One such polymer is cholestyramine, acopolymer of divinylbenzene/styrene and trimethylammonium methylstyrene.It has been long recognized that this polymer is unpalatable, gritty,and constipating. More recently, various polymers have been suggestedwhich are characterized by hydrophobic substituents and quaternaryammonium radicals substituted upon an amine polymer backbone (Ahlers, etal. U.S. Pat. Nos. 5,428,112 and 5,430,110 and McTaggert, et al., U.S.Pat. No. 5,462,730, which are incorporated herein by reference).

Thus, there is still a need to discover superior bile acid sequestrants.

SUMMARY OF THE INVENTION

The invention relates to the unexpected discovery that a new class ofion exchange resins have improved bile salt sequestration propertiesresulting in reduced dosages, which improve patient tolerance andcompliance, thereby improving the palatability of the composition andare relatively easy to manufacture. The polymers, employed in theinvention comprise non-absorbable, and optionally cross-linkedpolyamines as defined herein. The properties of the polymer which gaverise to the present invention were discovered during clinical trials ofthe polymer for its use in binding phosphate in patients suffering fromhyperphosphatemia. The polyamines of the invention are characterized byone or more monomeric units of the formula:

and salts thereof, where n is a positive integer and x is 0 or aninteger between 1 and about 4. The polymer can be characterized by thesubstantial absence of one or more alkylated amine monomers and/or thesubstantial absence of one or-more trialkylammonium alkyl groups. Inpreferred embodiments, the polymer is crosslinked by means of amultifunctional crosslinking agent.

The invention provides an effective treatment for removing bile saltsfrom a patient (and thereby reducing the patient's cholesterol level),particularly in patients with a serum LDL level of at least about 130mg/dL. The invention also provides for the use of the polymers describedherein for the manufacture of a medicament for the treatment ofhypercholesterolemia or for bile acid sequestration.

Other features and advantages will be apparent from the followingdescription of the preferred embodiments thereof and from the claims.

BRIEF DESCRIPTION OF THE DRAWING.

The Figure presents the effect of cross-linked polyallylamine on LDLcholesterol relative to baseline LDL cholesterol.

DETAILED DESCRIPTION OF THE INVENTION

As described above, the polymers employed in the invention comprise,optionally cross-linked polyamines characterized by the formula above.Preferred polymers are polyallylamine or polyvinylamine. Importantly,the polymers can be characterized by the substantial absence ofsubstituted or unsubstituted alkyl substituents on the amino group ofthe monomer, such as obtained in the alkylation of an amine polymer.That is, the polymer can be characterized in that the polymer issubstantially free of alkylated amine monomers.

The polymer can be a homopolymer or a copolymer of one or moreamine-containing monomers or non-amine containing monomers. Wherecopolymers are manufactured with the monomer of the above formula, thecomonomers are preferably inert, non-toxic and/or possess bile acidsequestration properties. Examples-of suitable non-amine-containingmonomers include vinylalcohol, acrylic acid, acrylamide, andvinylformamide. Examples of amine containing monomers preferably includemonomers having the Formula 1 above.

Preferably, the monomers are aliphatic. Most preferably, the polymer isa homopolymer, such as a homopolyallylamine or homopolyvinylamine.

Preferably, the polymer is rendered water-insoluble by crosslinking. Thecross-linking agent can be characterized by functional groups whichreact with the amino group of the monomer. Alternatively, thecrosslinking group can be characterized by two ore more vinyl groupswhich undergo free radical-polymerization with the amine monomer.

Examples of suitable crosslinking agents include acryloyl chloride,epichlorohydrin, butanedioldiglycidyl ether, ethanedioldiglycidyl ether,and dimethyl succinate.

A preferred crosslinking agent is epichlorohydrin because of its highavailability and low cost. Epichlorohydrin is also advantageous becauseof it's low molecular weight and hydrophilic nature, maintaining thewater-swellability of the polyamine gel.

The level of crosslinking makes the polymers insoluble and substantiallyresistant to absorption and degradation, thereby limiting the activityof the polymer to the gastrointestinal tract. Thus, the compositions arenon-systemic in their activity and will lead to reduced side-effects inthe patient. Typically, the cross-linking agent is present in an amountfrom about 0.5-25% (more preferably about 2.5-20% and most preferably1-10%). by weight, based upon total weight of monomer plus crosslinkingagent.

Typically, the amount of crosslinking-agent that is reacted with theamine polymer is sufficient to cause between about 0.5 and twentypercent of the amines. In a preferred embodiment, between about 0.5 and20 percent of the amine groups react with the crosslinking agent.

Preferred polymers of the invention are generally known in the art.Holmes-Farley, et al. (U.S. Pat. No. 5,496,545), describes the use ofaliphatic amine polymers in the treatment of hyperphosphatemia. Thesepolymers have also been suggested for use in the treatment ofiron-overload (Mandeville, et al., U.S. Pat. No. 5,487,888). Theteachings of both of these patents are incorporated herein by reference.

Non-cross-linked and cross-linked polyallylamine and polyvinylamine aregenerally known in the art and/or are commercially available. Methodsfor the manufacture of polyallylamine and polyvinylamine, andcross-linked derivatives thereof, are described in the above US Patents,the teachings of which are incorporated entirely by reference. Harada etal. (U.S. Pat. Nos. 4,605,701 and 4,528,347, which are incorporatedherein by reference in their entirety) also describe methods ofmanufacturing polyallylamine and cross-linked polyallylamine.

As described above the polymer can be administered in the form of asalt. By “salt” it is meant that the nitrogen group in the repeat unitis protonated to create a positively charged nitrogen atom associatedwith a negatively charged counterion.

The cationic counterions can be selected to minimize adverse effects onthe patient, as is more particularly described below. Examples ofsuitable counterions include Cl⁻, Br⁻, CH₃OSO₃ ⁻, HSO₄ ⁻, SO₄ ²⁻, HCO₃⁻, CO₃ ⁻, acetate, lactate, succinate, propionate, butyrate, ascorbate,citrate, maleate, folate, an amino acid derivative, a nucleotide, alipid, or a phospholipid. The counterions can be the same as, ordifferent from, each other. For example, the reaction product cancontain two different types of counterions, both of which are exchangedfor the bile salts being removed.

The polymers according to the invention can be administered orally to apatient in a dosage of about 1 mg/kg/day to about 1 g/kg/day, preferablybetween about 5 mg/kg/day to about 200 mg/kg/day (such as between about10 mg/kg/day to about 200 mg/kg/day); the particular dosage will dependon the individual-patient (e.g., the patient's weight and the extent ofbile salt removal required). The polymer can be administrated either inhydrated or dehydrated form, and can be flavored or added to a food ordrink, if desired to enhance patient acceptability. Additionalingredients such as other bile acid sequestrants, drugs for treatinghypercholesterolemia, atherosclerosis or other related indications, orinert ingredients, such as artificial coloring agents can be added aswell.

Examples of suitable forms for administration include tablets, capsules,and powders (e.g., for sprinkling on food) or mixing in water or juice).The tablet, capsule, or powder can be coated with a substance capable ofprotecting the composition from disintegration in the esophagus but willallow disintegration as the composition in the stomach and mixing withfood to pass into the patient's small intestine. The polymer can beadministered alone or in combination with a pharmaceutically acceptablecarrier substance, e.g., magnesium carbonate, lactose, or a phospholipidwith which the polymer can form a micelle.

The invention can be used to treat patients, preferably humans, withhypercholesterolemia, particularly patients with a serum LDL level whichexceeds about 130 mg/dL.

The invention will now be described more specifically by the examples.

EXAMPLES

A. Polymer Preparation

1. Preparation of Poly(vinylamine)

The first step involved the preparation of ethylidenebisacetamide.Acetamide (118 g), acetaldehyde (44.06 g), copper acetate (0.2 g), andwater (300 mL) were placed in a 1 L-three neck flask fitted withcondenser, thermometer, and mechanical stirred. Concentrated HCl (34 mL)was added and the mixture was heated to 45-50° C. with stirring for 24hours. The water was then removed in vacuo to leave a thick sludge whichformed crystals on cooling to 5° C. Acetone (200 mL) was added andstirred for a few minutes, after which the solid was filtered off anddiscarded. The acetone was cooled to 0° C. and solid was filtered off.This solid was rinsed in 500 mL acetone and air dried 18 hours to yield31.5 g of ethylidenebis-acetamide.

The next step involved the preparation of vinylacetamide fromethylidenebisacetamide. Ethylidenebisacetamide (31.05 g), calciumcarbonate (2 g) and celite 541 (2 g) were-placed in a 500 mL three neckflask fitted with a thermometer, a mechanical stirred, and a distillingheat atop a Vigroux column. The mixture was vacuum distilled at 24 mm Hgby heating the pot to 180-225° C. Only a single fraction was collected(10.8 g) which contained a large portion of acetamide in addition to theproduct (determined by NMR). This solid product was dissolved inisopropanol (30 mL) to form the crude vinylacetamide solution used forpolymerization.

Crude vinylacetamide solution (15 mL), divinylbenzene (1 g, technicalgrade, 55% pure, mixed isomers), and AIBN (0.3 g) were mixed and heatedto reflux under a nitrogen atmosphere for 90 minutes, forming a solidprecipitate. The solution was cooled, isopropanol. (50 mL) was added,and the solid was collected by centrifugation. The solid was rinsedtwice in isopropanol, once in water, and dried in a vacuum oven to yield0.8 g of poly(vinylacetamide), which was used to preparepoly(vinylamine).

Poly(vinylacetamide) (0.79 g) was placed in a 100 mL one neck flaskcontaining water (25 mL) and conc. HCl (25 mL). The mixture was refluxedfor 5 days, after which the solid was filtered of L, rinsed once inwater, twice in isopropanol, and dried in a vacuum oven to yield 0.77 gof product. Infrared spectroscopy indicated that a significant amount ofthe amide (1656 cm⁻¹) remained and that not much amine (1606 cm⁻¹) wasformed. The product of this reaction. (˜0.84 g) was suspended in NaOH(46 g) and water (46 g) and heated to boiling (˜140° C.). Due to foamingthe temperature was reduced and maintained at ˜100° C. for 2 hours.Water (10.0 mL) was added and the solid collected by filtration. Afterrinsing once in water the solid was suspended in water (500 mL) andadjusted to pH 5 with acetic acid. The solid was again filtered off,rinsed with water, then isopropanol, and dried in a vacuum oven to yield0.51 g of product. Infrared spectroscopy indicated that significantamine-had been formed.

2. Preparation of Poly(allylamine) Hydrochloride

To a 2 liter, water-jacketed reaction kettle equipped with (1) acondenser topped with a nitrogen gas inlet, (2) a thermometer, and (3) amechanical stirrer was added concentrated hydrochloric acid (360 mL).The acid was cooled to 5° C. using circulating water in the jacket ofthe reaction kettle (water temperature=0° C.). Allylamine (328.5 mL, 250g) was added dropwise with stirring while maintaining the reactiontemperature at 5-10° C. After addition was complete, the mixture wasremoved, placed in a 3 liter one-neck flask, and 206 g of liquid wasremoved by rotary vacuum evaporation at 60° C. Water (20 mL) was thenadded and the liquid was returned to the reaction kettle.Azobis(amidinopropane) dihydrochloride (0.5 g) suspended in 11 mL ofwater was then added. The resulting reaction mixture was heated to 50°C. under a nitrogen atmosphere with stirring for 24 hours. Additionalazobis(amidinopropane) dihydrochloride. (5 mL) suspended in 11 mL ofwater was then added, after which heating and stirring were continuedfor an additional 44 hours.

At the end of this period, distilled water (100 mL) was added to thereaction mixture and the liquid mixture allowed to cool with stirring.The mixture was then removed and placed in a 2 liter separatory funnel,after which it was added dropwise to a stirring solution of methanol (4L), causing a solid to form. The solid was removed by filtration,re-suspended in methanol (4 L), stirred for 1 hour, and collected byfiltration. The methanol rinse was then repeated one more time and thesolid dried in a vacuum oven to afford 215.1 g of poly(allylamine)hydrochloride as a granular white solid.

3. Preparation of Poly(allylamine) Hydrochloride Crosslinked withEpichlorohydrin

To a 5 gallon vessel was added poly(allylamine) hydrochloride preparedas described in Example 2 (1 kg) and water (4 L). The mixture wasstirred to dissolve the hydrochloride and the pH was adjusted by addingsolid NaOH (284 g). The resulting solution was cooled to roomtemperature, after which epichlorohydrin crosslinking agent (50 mL) wasadded all at once with stirring. The resulting mixture was stirredgently until it gelled (about 35 minutes). The crosslinking reaction wasallowed to proceed for an additional 18 hours at room temperature, afterwhich the polymer gel was removed and placed in portions in a blenderwith a total of 10 L of water. Each portion was blended gently for about3 minutes to form coarse particles which were then stirred for 1 hourand collected by filtration. The solid was rinsed three times bysuspending it in water (10 L, 15 L, 20 L), stirring each suspension for1 hour, and collecting the solid each time by filtration. The resultingsolid was then rinsed once by suspending it in isopropanol (17 L),stirring the mixture for 1 hour, and then collecting the solid byfiltration, after which the solid was dried in a vacuum oven at 50° C.for 18 hours to yield about 677 g of the cross linked polymer as agranular, brittle, white solid.

4. Preparation of Poly(allylamine) Hydrochloride Crosslinked withButanedioldiglycidyl Ether

To a 5 gallon vessle was added poly(allylamine) hydrochloride preparedas described in Example 2 (500 g) and water (2 L). The mixture wasstirred to dissolve the hydrochloride and the pH was adjusted to 10 byadding solid NaOH (134.6 g). The resulting solution was cooled to roomtemperature in the vessel, after which 1,4-butanedioldiglycidyl ethercrosslinking agent (65 mL) was added all at once with stirring. Theresulting mixture was stirred gently until it gelled (about 6 minutes).The crosslinking reaction was allowed to proceed for an additional 18hours at room temperature, after which the polymer gel was removed anddried in a vacuum oven at 75° C. for 24 hours. The dry solid was thenground and sieved to −30 mesh, after which it was suspended in 6 gallonsof water and stirred for 1 hour. The solid was then filtered off and therinse process repeated two more times. The resulting solid was then airdried for 4-8 hours, followed by drying in a vacuum oven at 50° C. for24 hours to yield about 415 g of the crosslinked polymer as a whitesolid.

5. Preparation of Poly(allylamine) Hydrochloride Crosslinked withEthanedioldiglycidyl Ether

To a 100 mL beaker was added poly(allylamine) hydrochloride prepared asdescribed in Example 2 (10 g) and water (40 mL). The mixture was stirredto dissolve the hydrochloride and the pH was adjusted to 10 by addingsolid NaOH. The resulting solution was cooled to room temperature in thebeaker, after which 1,2-ethanedioldiglycidyl ether crosslinking agent(2.0 mL) was added all at once with stirring. The resulting mixture wasstirred gently until it gelled (about 4 minutes). The crosslinkingreaction was allowed to proceed for an additional 18 hours at roomtemperature, after which the polymer gel was removed and blended in 500mL of methanol. The solid was then filtered off and suspended in water(500 mL). After stirring for 1 hour, the solid was filtered off and therinse process repeated. The resulting solid was rinsed twice inisopropanol (400 mL) and then dried-in a vacuum oven at 50° C. for 24hours to yield 8.7 g of the crosslinked polymer as a white solid.

6. Preparation of Poly(allylamine) Hydrochloride Crosslinked withDimethylsuccinate

To a 500 mL round-bottomed flask was added poly(allylamine)hydrochloride prepared as described in Example 2 (10 g), methanol (100mL), and triethylamine (10 mL). The mixture was stirred anddimethylsuccinate crosslinking agent (1 mL) was added. The solution washeated to reflux and the stirring discontinued after 30 minutes. After18 hours, the solution was cooled to room temperature, and the solidfiltered off and blended in 400 mL of isopropanol. The solid was thenfiltered off and suspended in water (1 L). After stirring for 1 hour,the solid was filtered off and the rinse process repeated two moretimes. The solid was then rinsed once in isopropanol (800 mL) and driedin a vacuum oven at 50° C. for 24 hours to yield 5.9 g of thecrosslinked polymer as a white solid.

An aqueous solution of poly(allylamine hydrochloride) (550 lb of a 50.7%aqueous solution) was diluted with water (751 lb) and neutralized withaqueous sodium hydroxide (171 lb of a 50% aqueous solution). Thesolution was cooled to approximately 25° C. and acetonitrile (1340 lb)and epichlorohydrin (26.2 lb) were added. The solution was stirredvigorously for 21 hours. During this time, the reactor contents changedfrom two liquid phases to a slurry, of particles in a liquid. The solidgel product was isolated by filtration. The gel was washed in anelutriation process with water (136,708 lb). The gel was isolated byfiltration and rinsed with isopropanol. The gel was slurried withisopropanol (1269 lb) and isolated by filtration. The isopropanol/waterwet gel was dried in a vacuum dryer at 60° C. The dried product wasground to pass through a 50 mesh screen to give a product suitable forpharmacologic use (166 lb, 73w).

7. Effect on Serum Cholesterol Levels in Humans

Hemodialysis patients on stable doses of calcium and/or aluminum basedphosphate binders entered a one-week screening period. The phosphatebinders were discontinued.

Those patients developing hyperphosphatemia (serum P04>6.0 mg/dL) duringthe wash-out period were eligible for drug treatment. A RenaGel® binder(epichlorohydrin cross-linked polyallylamine, GelTex Pharmaceuticals,Inc., Waltham, Mass.) starting dose was based on the degree ofhyperphosphatemia. Starting doses were either two, three, or four 465 mgcapsules three times per day with meals. At the end of each of threesubsequent two week periods, the dose of RenaGel® binder was increasedby one capsule per meal as necessary to achieve a serum phosphorusbetween 2.5 and 5.5 mg/dL, inclusive. If the serum phosphorus fell toless than 2.5 mg/dL, the RenaGel® binder dose was decreased by one tothree capsules per day to elevate the serum phosphorus to above 2.5mg/dL.

When the serum calcium fell below normal (defined by the centrallaboratory normal range) during the study, the serum calcium level wasreturned to within the normal range by adding an evening calciumsupplement of up to 1,000 mg of elemental calcium as the carbonate salton an empty stomach at bedtime or the dialysate calcium concentrationwas increased. TUMS EX® 750 mg tablets containing 300 mg of elementalcalcium were provided. Other brands of calcium carbonate or calciumacetate were used if the patient prefered another formulation.

At the conclusion of the treatment period, any remaining RenaGel®capsules were retrieved and the patient was kept off phosphate binderfor two weeks. After this second wash-out period, patients discontinuedany evening calcium supplements and returned to their original phosphatebinders.

Weekly throughout this period, on Mondays (MWF patients) and Tuesdays(TTS patients), the patients gave blood for the laboratory studies justprior to dialysis. On the Wednesdays (MWF patients) and Thursdays (TTSpatients) of the same weeks, the investigator inquired if the patientexperienced any adverse events or had changes in medications that mightindicate adverse events and reviewed the results of the laboratorytests.

Dietary intakes of phosphorus were assessed on selected days in thefirst wash-out, treatment, and second wash-out periods by 24-hour recallmethods by nutritionists from the University of Massachusetts MedicalCenter

Approximately 216 hemodialysis patients on stable doses of phosphatebinders were entered into the study. The patients had to have wellcontrolled serum phosphorus and not have any clinically significantunstable medical conditions. Only those patients who werehyperphosphatemiic (serum P04<6.0 mg/dL) during the first washout period(approximately 180 patients) received treatment.

The polymer was supplied as capsules containing 500 mg of polymer. Eachpatient started on one of three doses of polymer: (i) 2 capsules (0.93g) three times per day with meals; (ii) 3 capsules (1.4 g) three timesper day with meals; and (iii) 4 capsules (1.86 g) three times per daywith meals. Dose Level*** Overall Low Medium High Std P- Std Std Std P-Parameter Visit N Mean Dev Value* N Mean Dev N Mean Dev N Mean DevValue** Total Cholesterol −1 28 214.6 41.2 13 217.0 42.4 3 267.3 57.4 12198.8 23.8 0.0978 (mg/dL) 2 29 221.7 35.6 13 216.5 35.0 4 261.8 46.1 12214.0 25.1 0.0790 6 28 182.2 46.2 12 186.8 44.1 4 234.8 63.1 12 160.125.6 0.0222 10 25 184.7 48.5 12 195.5 47.7 4 223.5 52.9 9 153.1 29.00.0181 10/Final 25 184.7 48.5 12 195.5 47.7 4 223.5 52.9 9 153.1 29.00.0181 Change(10/Final − 2) 25 −37.2 29.0 <0.0001 12 −22.3 27.3 4 −38.325.3 9 −56.7 22.3 0.0098 12 25 208.1 42.1 12 202.6 38.4 4 267.3 45.6 9189.2 18.0 0.0291 Change(12 − 10) 24 23.1 34.2 0.0006 12 7.1 40.7 4 43.812.9 8 36.8 16.2 0.0306 LDL Cholesterol −1 27 145.0 34.1 12 147.2 32.2 3191.1 40.2 12 131.2 24.9 0.0494 (mg/dL) 2 29 154.6 27.4 13 147.4 16.3 4184.6 46.2 12 152.3 25.3 0.1441 6 28 110.5 33.4 12 113.3 32.4 4 150.543.9 12 94.5 17.3 0.0085 10 25 109.0 37.7 12 109.5 34.6 4 141.0 45.6 994.2 32.7 0.1750 10/Final 25 109.0 37.7 12 109.5 34.6 4 141.0 45.6 994.2 32.7 0.1750 Change(10/Final − 2) 25 −45.7 29.3 <0.0001 12 −38.029.0 4 −43.6 28.0 9 −56.8 29.9 0.2972 12 25 141.0 33.6 12 132.3 20.9 4194.2 37.9 9 129.0 23.8 0.0221 Change(12 − 10) 24 33.0 24.8 <0.0001 1222.8 23.6 4 53.2 17.9 8 38.2 23.9 0.0503 HDL Cholesterol −1 27 37.6 9.412 39.6 10.1 3 32.7 4.7 12 36.8 9.6 0.5108 (mg/dL) 2 29 36.4 9.2 13 37.89.8 4 31.3 5.0 12 36.5 9.6 0.4077 6 28 38.5 10.5 12 40.3 13.1 4 37.0 7.412 37.3 8.6 0.6622 10 25 36.5 11.1 12 41.3 12.0 4 34.5 6.1 9 30.9 9.30.1053 10/Final 25 36.5 11.1 12 41.3 12.0 4 34.5 6.1 9 30.9 9.3 0.1053Change(10/Final − 2) 25 0.8 9.0 0.2823 12 2.8 10.3 4 3.3 3.0 9 −3.0 8.20.1000 12 25 38.6 11.3 12 42.0 10.1 4 35.5 5.3 9 35.6 14.2 0.1986Change(12 − 10) 24 0.9 8.5 0.8018 12 0.7 7.7 4 1.0 2.7 8 1.3 11.8 0.7914Triglycerides −1 28 165.8 80.5 13 164.7 93.9 3 217.7 113.0 12 153.9 55.30.5796 (mg/dL) 2 29 153.9 92.3 13 156.3 103.7 4 229.5 104.0 12 126.264.0 0.2165 6 28 165.5 89.5 12 165.7 80.8 4 236.5 123.4 12 141.7 80.70.2408 10 25 196.2 165.3 12 223.4 222.6 4 240.0 65.1 9 140.3 81.8 0.099410/Final 25 196.2 165.3 12 223.4 222.6 4 240.0 65.1 9 140.3 81.8 0.0994Change(10/Final − 2) 25 38.2 150.6 0.3161 12 64.3 214.4 4 10.5 55.2 915.8 41.0 0.9199 12 25 142.5 91.2 12 141.7 107.2 4 188.0 76.3 9 123.474.3 0.2964 Change(12 − 10) 24 −54.0 151.3 0.0135 12 −81.8 209.6 4 −52.034.7 8 −13.4 49.7 0.2320*Wilcoxon Signed Rank Test**Kruskal-Wallis Exact Test***Dose level defined using the last actual dose during study8. Effect in Healthy Young and Old, Male and Female Volunteers

Eight young (19-40 years of age) and eight old (65 years of age andolder) healthy volunteer male and female subjects received 2.325 gramsof RenaGel® binder (epichlorohydrin cross-linked polyallylamine) threetimes per day with meals for 32 days. All drug doses were administeredwith meals served at a clinical research center for the entire 32 daystudy. On day 0, a 10 mL blood sample was drawn prior to the morningmeal and analyzed for plasma cholesterol levels. On day 32 a second 10mL blood sample was drawn prior to the morning meal. Subjects werereleased from the study after the morning meal on day 32. Plasmatriglycerides and HDL were measured and LDL cholesterol was calculatedby the Friedewald formula.

The Figure presents the effect of the the polymer on LDL cholesterolrelative to baseline LDL cholesterol. The higher the baselinecholesterol in these normal volunteers, the greater the decline in LDLcholesterol. LDL cholesterol declined by a mean of 42 mg/dL for theentire 16 patient cohort. Five patients in the study had baseline LDLcholesterol lower than 100 mg/dL. The decline in LDL cholesterol in the11 patients with baseline LDL cholesterol>than 120 mg/dL was 52.5 mg/dL.

EQUIVALENTS

Those skilled in the art will know, or be able to ascertain, using nomore than routine experimentation, many equivalents to the specificembodiments of the invention described herein. These and all otherequivalents are intended to be encompassed by the following claims.

1-15. (canceled)
 16. A method of removing bile salts from a patientcomprising administering to said patient a therapeutically effectiveamount of cross-linked homopolyallylamine, wherein said cross-linkedhomopolyallylamine is cross-linked by a multifunctional crosslinkingagent, and said multifunctional crosslinking agent is present in anamount from 0.5-25% by weight, based upon the combined weight ofallylamine repeat units and multifunctional crosslinking agent.
 17. Themethod of claim 16 wherein said multifunctional crosslinking agent ispresent in an amount from about 2.5-20% by weight, based upon thecombined weight of allylamine repeat units and multifunctionalcrosslinking agent.
 18. The method of claim 16, wherein the patient hasa serum LDL level of at least 130 mg/dL.
 19. The method of claim 16,wherein the multifunctional crosslinking agent comprisesepichlorohydrin.
 20. The method of claim 16, wherein the multifunctionalcrosslinking Agent is epichlorohydrin.